The separation of the various forms of rat liver microsomal cytochrome P450 will be pursued. Difference spectroscopy will be used as a refined tool to distinguish multiple forms. Reconstitution of P450 cytochromes into liposomes of different composition, along with varying quantities of P450-reductase and epoxide hydratase will be studied in detail. Benzo(a)pyrene metabolism and conjugation with DNA will be examined in these systems and will be compared to the same processes in microsomes and intact cells. The process of reduction of polycyclic hydrocarbon oxides will be characterized in detail with particular emphasis on the physiological relevance of the reaction. DNA modification by BP-metabolites will be examined with reference to the stability of the intermediates in cell membranes as compared to aqueous media. This will be extended to other unstable electrophilic species which are generated in microsomes or nuclear envelope and ultimately combined with DNA.